1 1906 140 ENHANCER OF ZESTE HOMOLOG 2-CATALYSED H3K27 TRIMETHYLATION PLAYS A KEY ROLE IN ACUTE-ON-CHRONIC LIVER FAILURE VIA TNF-MEDIATED PATHWAY. ACUTE-ON-CHRONIC LIVER FAILURE IS MAINLY DUE TO HOST IMMUNITY SELF-DESTRUCTION. THE HISTONE H3 LYSINE 27 (H3K27) TRIMETHYLATING ENZYME, ENHANCER OF ZESTE HOMOLOG 2 (EZH2) MEDIATES EPIGENETIC SILENCING OF GENE EXPRESSION AND REGULATES IMMUNITY, ALSO INVOLVES PATHOGENESIS OF SEVERAL LIVER DISEASES. THE CURRENT STUDY WAS TO DETERMINE THE ROLE OF METHYLTRANSFERASE EZH2 AND ITS CATALYSED H3K27 TRIMETHYLATION (H3K27ME3) IN LIVER FAILURE, AND TO FURTHER INVESTIGATE THE POTENTIAL TARGET FOR LIVER FAILURE TREATMENT. EZH2 AND ITS CATALYSED H3K27ME3 WERE DETERMINED IN PERIPHERAL BLOOD MONONUCLEAR CELLS (PBMC) FROM LIVER FAILURE PATIENTS AND KUPFFER CELLS FROM EXPERIMENTAL MICE. FURTHERMORE, GSK126 (AN INHIBITOR FOR EZH2 TRIMETHYLATION FUNCTION) WAS APPLIED IN LIVER FAILURE MICE IN VIVO, AND LIPOPOLYSACCHARIDE-STIMULATED MONONUCLEAR CELLS IN VITRO. EZH2 AND H3K27ME3 WERE SIGNIFICANTLY UPREGULATED IN HUMAN PBMC FROM LIVER FAILURE PATIENTS OR MURINE KUPFFER CELLS FROM THE LIVER FAILURE ANIMALS, RESPECTIVELY. GSK126 AMELIORATED DISEASE SEVERITY IN LIVER FAILURE MICE, WHICH MAYBE ATTRIBUTE TO DOWN-REGULATE CIRCULATING AND HEPATIC PROINFLAMMATORY CYTOKINES, ESPECIALLY TNF VIA REDUCING H3K27ME3. IN-DEPTH CHROMATIN IMMUNOPRECIPITATION ANALYSIS UNRAVELLED THAT DECREASED ENRICHMENT OF H3K27ME3 ON TNF PROMOTOR, RESULTING IN TNF ELEVATION IN KUPFFER CELLS FROM LIVER FAILURE MICE. NUCLEAR FACTOR KAPPA B (NF-KAPPAB) AND PROTEIN KINASE B (AKT) SIGNALLING PATHWAYS WERE ACTIVATED UPON LIPOPOLYSACCHARIDE STIMULATION, BUT ATTENUATED BY USING GSK126, ACCOMPANIED WITH DECREASED TNF IN VITRO. IN CONCLUSION, EZH2 AND H3K27ME3 CONTRIBUTED TO THE PATHOGENESIS OF LIVER FAILURE VIA TRIGGERING TNF AND OTHER INDISPENSABLE PROINFLAMMATORY CYTOKINES. EZH2 WAS TO MODIFY H3K27ME3 ENRICHMENT, AS WELL AS, ACTIVATION OF THE DOWNSTREAM NF-KAPPAB AND AKT SIGNALLING PATHWAYS. 2018 2 2825 53 FLOW-DEPENDENT EPIGENETIC REGULATION OF IGFBP5 EXPRESSION BY H3K27ME3 CONTRIBUTES TO ENDOTHELIAL ANTI-INFLAMMATORY EFFECTS. RATIONALE: ATHEROSCLEROSIS IS A CHRONIC INFLAMMATORY AND EPIGENETIC DISEASE THAT IS INFLUENCED BY DIFFERENT PATTERNS OF BLOOD FLOW. HOWEVER, THE EPIGENETIC MECHANISM WHEREBY ATHEROPROTECTIVE FLOW CONTROLS ENDOTHELIAL GENE PROGRAMMING REMAINS ELUSIVE. HERE, WE INVESTIGATED THE POSSIBILITY THAT FLOW ALTERS ENDOTHELIAL GENE EXPRESSION THROUGH EPIGENETIC MECHANISMS. METHODS: EN FACE STAINING AND WESTERN BLOT WERE USED TO DETECT PROTEIN EXPRESSION. REAL-TIME PCR WAS USED TO DETERMINE RELATIVE GENE EXPRESSION. RNA-SEQUENCING OF HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS TREATED WITH SIRNA OF ENHANCER OF ZESTE HOMOLOG 2 (EZH2) OR LAMINAR FLOW WAS USED FOR TRANSCRIPTIONAL PROFILING. RESULTS: WE FOUND THAT TRIMETHYLATION OF HISTONE 3 LYSINE 27 (H3K27ME3), A REPRESSIVE EPIGENETIC MARK THAT ORCHESTRATES GENE REPRESSION, WAS REDUCED IN LAMINAR FLOW AREAS OF MOUSE AORTA AND FLOW-TREATED HUMAN ENDOTHELIAL CELLS. THE DECREASE OF H3K27ME3 PARALLELED A REDUCTION IN THE EPIGENETIC "WRITER"-EZH2, THE CATALYTIC SUBUNIT OF THE POLYCOMB REPRESSIVE COMPLEX 2 (PRC2). MOREOVER, LAMINAR FLOW DECREASED EXPRESSION OF EZH2 VIA MECHANOSENSITIVE MIR101. GENOME-WIDE TRANSCRIPTOME PROFILING STUDIES IN ENDOTHELIAL CELLS TREATED WITH EZH2 SIRNA AND FLOW REVEALED THE UPREGULATION OF NOVEL MECHANOSENSITIVE GENE IGFBP5 (INSULIN-LIKE GROWTH FACTOR-BINDING PROTEIN 5), WHICH IS EPIGENETICALLY SILENCED BY H3K27ME3. FUNCTIONALLY, INHIBITION OF H3K27ME3 BY EZH2 SIRNA OR GSK126 (A SPECIFIC EZH2 INHIBITOR) REDUCED H3K27ME3 LEVELS AND MONOCYTE ADHESION TO ENDOTHELIAL CELLS. ADENOVIRAL OVEREXPRESSION OF IGFBP5 ALSO RECAPITULATED THE ANTI-INFLAMMATORY EFFECTS OF H3K27ME3 INHIBITION. MORE IMPORTANTLY, WE OBSERVED EZH2 UPREGULATION, AND IGFBP5 DOWNREGULATION, IN ADVANCED ATHEROSCLEROTIC PLAQUES FROM HUMAN PATIENTS. CONCLUSION: TAKEN TOGETHER, OUR FINDINGS REVEAL THAT ATHEROPROTECTIVE FLOW REDUCES H3K27ME3 AS A CHROMATIN-BASED MECHANISM TO AUGMENT THE EXPRESSION OF GENES THAT CONFER AN ANTI-INFLAMMATORY RESPONSE IN THE ENDOTHELIUM. OUR STUDY EXEMPLIFIES FLOW-DEPENDENT EPIGENETIC REGULATION OF ENDOTHELIAL GENE EXPRESSION, AND ALSO SUGGESTS THAT TARGETING THE EZH2/H3K27ME3/IGFBP5 PATHWAY MAY OFFER NOVEL THERAPEUTICS FOR INFLAMMATORY DISORDERS SUCH AS ATHEROSCLEROSIS. 2018 3 4546 33 MUTANT P53 REGULATES ENHANCER-ASSOCIATED H3K4 MONOMETHYLATION THROUGH INTERACTIONS WITH THE METHYLTRANSFERASE MLL4. MONOMETHYLATION OF HISTONE H3 LYSINE 4 (H3K4ME1) IS ENRICHED AT ENHANCERS THAT ARE PRIMED FOR ACTIVATION AND THE LEVELS OF THIS HISTONE MARK ARE FREQUENTLY ALTERED IN VARIOUS HUMAN CANCERS. YET, HOW ALTERATIONS IN H3K4ME1 ARE ESTABLISHED AND THE CONSEQUENCES OF THESE EPIGENETIC CHANGES IN TUMORIGENESIS ARE NOT WELL UNDERSTOOD. USING CHIP-SEQ IN HUMAN COLON CANCER CELLS, WE DEMONSTRATE THAT MUTANT P53 DEPLETION RESULTS IN DECREASED H3K4ME1 LEVELS AT ACTIVE ENHANCERS THAT REVEAL A STRIKING COLOCALIZATION OF MUTANT P53 AND THE H3K4 MONOMETHYLTRANSFERASE MLL4 FOLLOWING CHRONIC TUMOR NECROSIS FACTOR ALPHA (TNFALPHA) SIGNALING. WE FURTHER REVEAL THAT MUTANT P53 FORMS PHYSIOLOGICAL ASSOCIATIONS AND DIRECT INTERACTIONS WITH MLL4 AND PROMOTES THE ENHANCER BINDING OF MLL4, WHICH IS REQUIRED FOR TNFALPHA-INDUCIBLE H3K4ME1 AND HISTONE H3 LYSINE 27 ACETYLATION (H3K27AC) LEVELS, ENHANCER-DERIVED TRANSCRIPT (ERNA) SYNTHESIS, AND MUTANT P53-DEPENDENT TARGET GENE ACTIVATION. COMPLEMENTARY IN VITRO STUDIES WITH RECOMBINANT CHROMATIN AND PURIFIED PROTEINS DEMONSTRATE THAT BINDING OF THE MLL3/4 COMPLEX AND H3K4ME1 DEPOSITION IS ENHANCED BY MUTANT P53 AND P300-MEDIATED ACETYLATION, WHICH IN TURN REFLECTS A MLL3/4-DEPENDENT ENHANCEMENT OF MUTANT P53 AND P300-DEPENDENT TRANSCRIPTIONAL ACTIVATION. COLLECTIVELY, OUR FINDINGS ESTABLISH A MECHANISM IN WHICH MUTANT P53 COOPERATES WITH MLL4 TO REGULATE ABERRANT ENHANCER ACTIVITY AND TUMOR-PROMOTING GENE EXPRESSION IN RESPONSE TO CHRONIC IMMUNE SIGNALING. 2018 4 2300 31 EPIGENETIC REGULATION OF BDNF EXPRESSION IN THE PRIMARY SENSORY NEURONS AFTER PERIPHERAL NERVE INJURY: IMPLICATIONS IN THE DEVELOPMENT OF NEUROPATHIC PAIN. BRAIN-DERIVED NEUROTROPHIC FACTOR (BDNF) IS KNOWN TO BE UP-REGULATED IN THE DORSAL ROOT GANGLION (DRG) AFTER PERIPHERAL NERVE INJURY, AND TO CONTRIBUTE TO NEUROPATHIC PAIN. HERE, WE FOUND THAT THERMAL HYPERALGESIA AND MECHANICAL ALLODYNIA AT DAY 7 POST-INJURY WERE INHIBITED ONLY WHEN ANTI-BDNF ANTIBODY WAS INTRATHECALLY ADMINISTRATED AT DAY 2 POST-INJURY. CONSISTENT WITH BEHAVIORAL RESULTS, WESTERN BLOT ANALYSIS SHOWED THAT THE EXPRESSION LEVELS OF BDNF PROTEIN IN THE SPINAL DORSAL HORN WERE MARKEDLY INDUCED DURING EARLY STAGE POST-INJURY. MOREOVER, THE MAXIMAL INCREASE IN BDNF MRNA EXPRESSION IN THE DRG WAS OBSERVED AT DAY 1 POST-INJURY, AND SIGNIFICANTLY ELEVATED LEVELS WERE SUSTAINED FOR AT LEAST 14 DAYS. FOUR OF FIVE BDNF MRNA TRANSCRIPTS WERE UP-REGULATED AFTER NERVE INJURY, AND THE MOST INDUCIBLE TRANSCRIPT WAS EXON I. USING A CHROMATIN IMMUNOPRECIPITATION (CHIP) ASSAY, WE FOUND THAT NERVE INJURY PROMOTES HISTONE H3 AND H4 ACETYLATION, TRANSCRIPTIONALLY ACTIVE MODIFICATIONS, AT BDNF PROMOTER I AT DAY 1 POST-INJURY, AND THE LEVELS OF HISTONE ACETYLATION REMAIN ELEVATED FOR AT LEAST 7 DAYS. TAKEN TOGETHER, OUR FINDINGS SUGGEST THAT AN INITIAL INCREASE IN BDNF EXON I EXPRESSION CONTROLLED BY EPIGENETIC MECHANISMS MIGHT HAVE A CRUCIAL ROLE IN THE DEVELOPMENT OF NEUROPATHIC PAIN. 2013 5 6660 34 UPREGULATION OF CXCR4 THROUGH PROMOTER DEMETHYLATION CONTRIBUTES TO INFLAMMATORY HYPERALGESIA IN RATS. AIM AND METHODS: CHRONIC PAIN ASSOCIATED WITH INFLAMMATION IS A COMMON CLINICAL PROBLEM, AND THE UNDERLYING MECHANISMS YET ARE INCOMPLETELY DEFINED. DNA METHYLATION HAS BEEN IMPLICATED IN THE PATHOGENESIS OF CHRONIC PAIN. HOWEVER, THE SPECIFIC GENES REGULATED BY DNA METHYLATION UNDER INFLAMMATORY PAIN CONDITION REMAIN LARGELY UNKNOWN. HERE, WE INVESTIGATED HOW CHEMOKINE RECEPTOR CXCR4 EXPRESSION IS REGULATED BY DNA METHYLATION AND HOW IT CONTRIBUTES TO INFLAMMATORY PAIN INDUCED BY COMPLETE FREUND'S ADJUVANT (CFA) IN RATS. RESULTS: INTRAPLANTAR INJECTION OF CFA COULD NOT ONLY INDUCE SIGNIFICANT HYPERALGESIA IN RATS, BUT ALSO SIGNIFICANTLY INCREASE THE EXPRESSION OF CXCR4 MRNA AND PROTEIN IN THE DORSAL ROOT GANGLION (DRG). INTRATHECAL INJECTION OF CXCR4 ANTAGONIST AMD3100 SIGNIFICANTLY RELIEVED HYPERALGESIA IN INFLAMMATORY RATS IN A TIME- AND DOSE-DEPENDENT MANNER. BISULFITE SEQUENCING AND METHYLATION-SPECIFIC PCR DEMONSTRATE THAT CFA INJECTION LED TO A SIGNIFICANT DEMETHYLATION OF CPG ISLAND AT CXCR4 GENE PROMOTER. CONSISTENTLY, THE EXPRESSION OF DNMT3B WAS SIGNIFICANTLY DOWNREGULATED AFTER CFA INJECTION. ONLINE SOFTWARE PREDICTION REVEALS THREE BINDING SITES OF P65 IN THE CPG ISLAND OF CXCR4 GENE PROMOTER, WHICH HAS CONFIRMED BY THE CHROMATIN IMMUNOPRECIPITATION ASSAY, CFA TREATMENT SIGNIFICANTLY INCREASES THE RECRUITMENT OF P65 TO CXCR4 GENE PROMOTER. INHIBITION OF NF-KB SIGNALING USING P65 INHIBITOR PYRROLIDINE DITHIOCARBAMATE SIGNIFICANTLY PREVENTED THE INCREASES OF THE CXCR4 EXPRESSION. CONCLUSION: UPREGULATION OF CXCR4 EXPRESSION DUE TO PROMOTER DEMETHYLATION FOLLOWED BY INCREASED RECRUITMENT OF P65 TO PROMOTER OF CXCR4 GENE CONTRIBUTES TO INFLAMMATORY HYPERALGESIA. THESE FINDINGS PROVIDE A THEORETICAL BASIS FOR THE TREATMENT OF CHRONIC PAIN FROM AN EPIGENETIC PERSPECTIVE. 2018 6 2785 39 EZH2 REGULATES SPINAL NEUROINFLAMMATION IN RATS WITH NEUROPATHIC PAIN. ALTERATION IN GENE EXPRESSION ALONG THE PAIN SIGNALING PATHWAY IS A KEY MECHANISM CONTRIBUTING TO THE GENESIS OF NEUROPATHIC PAIN. ACCUMULATING STUDIES HAVE SHOWN THAT EPIGENETIC REGULATION PLAYS A CRUCIAL ROLE IN NOCICEPTIVE PROCESS IN THE SPINAL DORSAL HORN. IN THIS PRESENT STUDY, WE INVESTIGATED THE ROLE OF ENHANCER OF ZESTE HOMOLOG-2 (EZH2), A SUBUNIT OF THE POLYCOMB REPRESSIVE COMPLEX 2, IN THE SPINAL DORSAL HORN IN THE GENESIS OF NEUROPATHIC PAIN IN RATS INDUCED BY PARTIAL SCIATIC NERVE LIGATION. EZH2 IS A HISTONE METHYLTRANSFERASE, WHICH CATALYZES THE METHYLATION OF HISTONE H3 ON K27 (H3K27), RESULTING IN GENE SILENCING. WE FOUND THAT LEVELS OF EZH2 AND TRI-METHYLATED H3K27 (H3K27TM) IN THE SPINAL DORSAL HORN WERE INCREASED IN RATS WITH NEUROPATHIC PAIN ON DAY 3 AND DAY 10 POST NERVE INJURIES. EZH2 WAS PREDOMINANTLY EXPRESSED IN NEURONS IN THE SPINAL DORSAL HORN UNDER NORMAL CONDITIONS. THE NUMBER OF NEURONS WITH EZH2 EXPRESSION WAS INCREASED AFTER NERVE INJURY. MORE STRIKINGLY, NERVE INJURY DRASTICALLY INCREASED THE NUMBER OF MICROGLIA WITH EZH2 EXPRESSION BY MORE THAN SEVENFOLD. INTRATHECAL INJECTION OF THE EZH2 INHIBITOR ATTENUATED THE DEVELOPMENT AND MAINTENANCE OF MECHANICAL AND THERMAL HYPERALGESIA IN RATS WITH NERVE INJURY. SUCH ANALGESIC EFFECTS WERE CONCURRENTLY ASSOCIATED WITH THE REDUCED LEVELS OF EZH2, H3K27TM, IBA1, GFAP, TNF-ALPHA, IL-1BETA, AND MCP-1 IN THE SPINAL DORSAL HORN IN RATS WITH NERVE INJURY. OUR RESULTS HIGHLY SUGGEST THAT TARGETING THE EZH2 SIGNALING PATHWAY COULD BE AN EFFECTIVE APPROACH FOR THE MANAGEMENT OF NEUROPATHIC PAIN. 2017 7 4906 40 P300 EXERTS AN EPIGENETIC ROLE IN CHRONIC NEUROPATHIC PAIN THROUGH ITS ACETYLTRANSFERASE ACTIVITY IN RATS FOLLOWING CHRONIC CONSTRICTION INJURY (CCI). BACKGROUND: NEUROPATHIC PAIN IS DETRIMENTAL TO HUMAN HEALTH; HOWEVER, ITS PATHOGENESIS STILL REMAINS LARGELY UNKNOWN. OVEREXPRESSION OF PAIN-ASSOCIATED GENES AND INCREASED NOCICEPTIVE SOMATO-SENSITIVITY ARE WELL OBSERVED IN NEUROPATHIC PAIN. THE IMPORTANCE OF EPIGENETIC MECHANISMS IN REGULATING THE EXPRESSION OF PRO- OR ANTI-NOCICEPTIVE GENES HAS BEEN REVEALED BY STUDIES RECENTLY, AND WE HYPOTHESIZE THAT THE TRANSCRIPTIONAL COACTIVATOR AND THE HISTONE ACETYLTRANSFERASE E1A BINDING PROTEIN P300 (P300), AS A PART OF THE EPIGENETIC MECHANISMS OF GENE REGULATION, MAY BE INVOLVED IN THE PATHOGENESIS OF NEUROPATHIC PAIN INDUCED BY CHRONIC CONSTRICTION INJURY (CCI). TO TEST THIS HYPOTHESIS, TWO DIFFERENT APPROACHES WERE USED IN THIS STUDY: (I) DOWN-REGULATING P300 WITH SPECIFIC SMALL HAIRPIN RNA (SHRNA) AND (II) CHEMICAL INHIBITION OF P300 ACETYLTRANSFERASE ACTIVITY BY A SMALL MOLECULE INHIBITOR, C646. RESULTS: USING THE CCI RAT MODEL, WE FOUND THAT THE P300 EXPRESSION WAS INCREASED IN THE LUMBAR SPINAL CORD ON DAY 14 AFTER CCI. THE TREATMENT WITH INTRATHECAL P300 SHRNA REVERSED CCI-INDUCED MECHANICAL ALLODYNIA AND THERMAL HYPERALGESIA, AND SUPPRESSED THE EXPRESSION OF CYCLOOXYGENASE-2 (COX-2), A NEUROPATHIC PAIN-ASSOCIATED FACTOR. FURTHERMORE, C646, AN INHIBITOR OF P300 ACETYLTRANSFERASE, ALSO ATTENUATED MECHANICAL ALLODYNIA AND THERMAL HYPERALGESIA, ACCOMPANIED BY A SUPPRESSED COX-2 EXPRESSION, IN THE SPINAL CORD. CONCLUSIONS: THE RESULTS SUGGEST THAT, THROUGH ITS ACETYLTRANSFERASE ACTIVITY IN THE SPINAL CORD AFTER CCI, P300 EPIGENETICALLY PLAYS AN IMPORTANT ROLE IN NEUROPATHIC PAIN. INHIBITING P300, USING INTERFERING RNA OR C646, MAY BE A PROMISING APPROACH TO THE DEVELOPMENT OF NEW NEUROPATHIC PAIN THERAPIES. 2012 8 1320 33 DEMETHYLATION REGULATION OF BDNF GENE EXPRESSION IN DORSAL ROOT GANGLION NEURONS IS IMPLICATED IN OPIOID-INDUCED PAIN HYPERSENSITIVITY IN RATS. REPEATED ADMINISTRATION OF MORPHINE MAY RESULT IN OPIOID-INDUCED HYPERSENSITIVITY (OIH), WHICH INVOLVES ALTERED EXPRESSION OF NUMEROUS GENES, INCLUDING BRAIN-DERIVED NEUROTROPHIC FACTOR (BDNF) IN DORSAL ROOT GANGLION (DRG) NEURONS. YET, IT REMAINS UNCLEAR HOW BDNF EXPRESSION IS INCREASED IN DRG NEURONS AFTER REPEATED MORPHINE TREATMENT. DNA METHYLATION IS AN IMPORTANT MECHANISM OF EPIGENETIC CONTROL OF GENE EXPRESSION. IN THE CURRENT STUDY, WE HYPOTHESIZED THAT THE DEMETHYLATION REGULATION OF CERTAIN BDNF GENE PROMOTERS IN DRG NEURONS MAY CONTRIBUTE TO THE DEVELOPMENT OF OIH. REAL-TIME RT-PCR WAS USED TO ASSESS CHANGES IN THE MRNA TRANSCRIPTION LEVELS OF MAJOR BDNF EXONS INCLUDING EXON I, II, IV, VI, AS WELL AS TOTAL BDNF MRNA IN DRGS FROM RATS AFTER REPEATED MORPHINE ADMINISTRATION. THE LEVELS OF EXON IV AND TOTAL BDNF MRNA WERE SIGNIFICANTLY UPREGULATED BY REPEATED MORPHINE ADMINISTRATION, AS COMPARED TO THAT IN SALINE CONTROL GROUP. FURTHER, ELISA ARRAY AND IMMUNOCYTOCHEMISTRY STUDY REVEALED A ROBUST UPREGULATION OF BDNF PROTEIN EXPRESSION IN DRG NEURONS AFTER REPEATED MORPHINE EXPOSURE. CORRESPONDINGLY, THE METHYLATION LEVELS OF BDNF EXON IV PROMOTER SHOWED A SIGNIFICANT DOWNREGULATION BY MORPHINE TREATMENT. IMPORTANTLY, INTRATHECAL ADMINISTRATION OF A BDNF ANTIBODY, BUT NOT CONTROL IGG, SIGNIFICANTLY INHIBITED MECHANICAL HYPERSENSITIVITY THAT DEVELOPED IN RATS AFTER REPEATED MORPHINE TREATMENT. CONVERSELY, INTRATHECAL ADMINISTRATION OF AN INHIBITOR OF DNA METHYLATION, 5-AZA-2'-DEOXYCYTIDINE (5-AZA-DC) MARKEDLY UPREGULATED THE BDNF PROTEIN EXPRESSION IN DRG NEURONS AND ENHANCED THE MECHANICAL ALLODYNIA AFTER REPEATED MORPHINE EXPOSURE. TOGETHER, OUR FINDINGS SUGGEST THAT DEMETHYLATION REGULATION OF BDNF GENE PROMOTER MAY BE IMPLICATED IN THE DEVELOPMENT OF OIH THROUGH EPIGENETIC CONTROL OF BDNF EXPRESSION IN DRG NEURONS. 2016 9 2783 44 EZH2 METHYLTRANSFERASE REGULATES NEUROINFLAMMATION AND NEUROPATHIC PAIN. RECENT STUDIES BY US AND OTHERS HAVE SHOWN THAT ENHANCER OF ZESTE HOMOLOG-2 (EZH2), A HISTONE METHYLTRANSFERASE, IN GLIAL CELLS REGULATES THE GENESIS OF NEUROPATHIC PAIN BY MODULATING THE PRODUCTION OF PROINFLAMMATORY CYTOKINES AND CHEMOKINES. IN THIS REVIEW, WE SUMMARIZE RECENT ADVANCES IN THIS RESEARCH AREA. EZH2 IS A SUBUNIT OF POLYCOMB REPRESSIVE COMPLEX 2 (PRC2), WHICH PRIMARILY SERVES AS A HISTONE METHYLTRANSFERASE TO CATALYZE METHYLATION OF HISTONE 3 ON LYSINE 27 (H3K27), ULTIMATELY RESULTING IN TRANSCRIPTIONAL REPRESSION. ANIMALS WITH NEUROPATHIC PAIN EXHIBIT INCREASED EZH2 ACTIVITY AND NEUROINFLAMMATION OF THE INJURED NERVE, SPINAL CORD, AND ANTERIOR CINGULATE CORTEX. INHIBITION OF EZH2 WITH DZNEP OR GSK-126 AMELIORATES NEUROINFLAMMATION AND NEUROPATHIC PAIN. EZH2 PROTEIN EXPRESSION INCREASES UPON ACTIVATION OF TOLL-LIKE RECEPTOR 4 AND CALCITONIN GENE-RELATED PEPTIDE RECEPTORS, DOWNREGULATION OF MIR-124-3P AND MIR-378 MICRORNAS, OR UPREGULATION OF LNCENC1 AND MALAT1 LONG NONCODING RNAS. GENES SUPPRESSED BY EZH2 INCLUDE SUPPRESSOR OF CYTOKINE SIGNALING 3 (SOCS3), NUCLEAR FACTOR (ERYTHROID-DERIVED 2)-LIKE-2 FACTOR (NRF2), MIR-29B-3P, MIR-146A-5P, AND BRAIN-SPECIFIC ANGIOGENESIS INHIBITOR 1 (BAI1). PRO-INFLAMMATORY MEDIATORS FACILITATE NEURONAL ACTIVATION ALONG PAIN-SIGNALING PATHWAYS BY SENSITIZING NOCICEPTORS IN THE PERIPHERY, AS WELL AS ENHANCING EXCITATORY SYNAPTIC ACTIVITIES AND SUPPRESSING INHIBITORY SYNAPTIC ACTIVITIES IN THE CNS. THESE STUDIES COLLECTIVELY REVEAL THAT EZH2 IS IMPLICATED IN SIGNALING PATHWAYS KNOWN TO BE KEY PLAYERS IN THE PROCESS OF NEUROINFLAMMATION AND GENESIS OF NEUROPATHIC PAIN. THEREFORE, TARGETING THE EZH2 SIGNALING PATHWAY MAY OPEN A NEW AVENUE TO MITIGATE NEUROINFLAMMATION AND NEUROPATHIC PAIN. 2023 10 717 47 CALCITONIN GENE-RELATED PEPTIDE REGULATES SPINAL MICROGLIAL ACTIVATION THROUGH THE HISTONE H3 LYSINE 27 TRIMETHYLATION VIA ENHANCER OF ZESTE HOMOLOG-2 IN RATS WITH NEUROPATHIC PAIN. BACKGROUND: CALCITONIN GENE-RELATED PEPTIDE (CGRP) AS A MEDIATOR OF MICROGLIAL ACTIVATION AT THE TRANSCRIPTIONAL LEVEL MAY FACILITATE NOCICEPTIVE SIGNALING. TRIMETHYLATION OF H3 LYSINE 27 (H3K27ME3) BY ENHANCER OF ZESTE HOMOLOG 2 (EZH2) IS AN EPIGENETIC MARK THAT REGULATES INFLAMMATORY-RELATED GENE EXPRESSION AFTER PERIPHERAL NERVE INJURY. IN THIS STUDY, WE EXPLORED THE RELATIONSHIP BETWEEN CGRP AND H3K27ME3 IN MICROGLIAL ACTIVATION AFTER NERVE INJURY, AND ELUCIDATED THE UNDERLYING MECHANISMS IN THE PATHOGENESIS OF CHRONIC NEUROPATHIC PAIN. METHODS: MICROGLIAL CELLS (BV2) WERE TREATED WITH CGRP AND DIFFERENTIALLY ENRICHMENTS OF H3K27ME3 ON GENE PROMOTERS WERE EXAMINED USING CHIP-SEQ. A CHRONIC CONSTRICTION INJURY (CCI) RAT MODEL WAS USED TO EVALUATE THE ROLE OF CGRP ON MICROGLIAL ACTIVATION AND EZH2/H3K27ME3 SIGNALING IN CCI-INDUCED NEUROPATHIC PAIN. RESULTS: OVEREXPRESSIONS OF EZH2 AND H3K27ME3 WERE CONFIRMED IN SPINAL MICROGLIA OF CCI RATS BY IMMUNOFLUORESCENCE. CGRP TREATMENT INDUCED THE INCREASED OF H3K27ME3 EXPRESSION IN THE SPINAL DORSAL HORN AND CULTURED MICROGLIAL CELLS (BV2) THROUGH EZH2. CHIP-SEQ DATA INDICATED THAT CGRP SIGNIFICANTLY ALTERED H3K27ME3 ENRICHMENTS ON GENE PROMOTERS IN MICROGLIA FOLLOWING CGRP TREATMENT, INCLUDING 173 GAINING H3K27ME3 AND 75 LOSING THIS MARK, WHICH MOSTLY ENRICHED IN REGULATION OF CELL GROWTH, PHAGOSOME, AND INFLAMMATION. QRT-PCR VERIFIED EXPRESSIONS OF REPRESENTATIVE CANDIDATE GENES (TRAF3IP2, BCL2L11, ITGAM, DAB2, NLRP12, WNT3, ADAM10) AND REAL-TIME CELL ANALYSIS (RTCA) VERIFIED MICROGLIAL PROLIFERATION. ADDITIONALLY, CGRP TREATMENT AND CCI INCREASED EXPRESSIONS OF ITGAM, ADAM10, MCP-1, AND CX3CR1, KEY MEDIATORS OF MICROGLIAL ACTIVATION IN SPINAL DORSAL HORN AND CULTURED MICROGLIAL CELLS. SUCH INCREASED EFFECTS INDUCED BY CCI WERE SUPPRESSED BY CGRP ANTAGONIST AND EZH2 INHIBITOR, WHICH WERE CONCURRENTLY ASSOCIATED WITH THE ATTENUATED MECHANICAL AND THERMAL HYPERALGESIA IN CCI RATS. CONCLUSION: OUR FINDINGS HIGHLY INDICATE THAT CGRP IS IMPLICATED IN THE GENESIS OF NEUROPATHIC PAIN THROUGH REGULATING MICROGLIAL ACTIVATION VIA EZH2-MEDIATED H3K27ME3 IN THE SPINAL DORSAL HORN. 2021 11 3832 29 INVOLVEMENT OF SPINAL SIRT1 IN DEVELOPMENT OF CHRONIC CONSTRICTION INJURY INDUCED NEUROPATHIC PAIN IN RATS. IT IS KNOWN THAT THE EPIGENETIC PROCESS OF HISTONE ACETYLATION IS INVOLVED IN THE NEUROPATHIC PAIN. THE AIM OF THIS STUDY WAS TO DETERMINE WHETHER SIRTUIN TYPE 1 (SIRT1), AN NAD(+) DEPENDENT DEACETYLASE, AFFECTED ALLODYNIA AND HYPERALGESIA IN NEUROPATHIC PAIN. THE NEUROPATHIC PAIN MODEL WAS ESTABLISHED BY LIGATURE OF THE RIGHT SCIATIC NERVE TO INDUCE CHRONIC CONSTRICTION INJURY (CCI) IN RATS. HISTONE ACETYLTRANSFERASE (HAT) ACTIVITY WAS INCREASED AND, AND HISTONE DEACETYLASE (HDAC) ACTIVITY WAS DECLINED IN TISSUE OF THE SPINAL DORSA HORN IN CCI RATES BY MEANS OF ENZYME-LINKED IMMUNOSORBENT ASSAY (ELISA). THE PERSISTENT HYPERALGESIA AND ALLODYNIA CAUSED BY CCI WERE ASSOCIATED WITH DOWNREGULATION OF SIRT1 AND UPREGULATION OF ACETYLATED-H3 (AC-H3) IN TISSUE OF THE SPINAL CORD BY WESTERN BLOT ASSAY, WHICH WAS REVERSED AFTER INTRATHECAL INJECTION OF SIRT1 AGONIST SRT1720. SRT1720 TREATMENT ACHIEVED ANALGESIC THROUGH INHIBITING THE ACETYLATION OF NUCLEAR FACTOR KAPPA B (NF-KAPPAB) AND BLOCKING THE RELEASES OF THE INFLAMMATORY FACTORS INCLUDING TUMOR NECROSIS FACTOR-ALPHA (TNF-ALPHA) AND INTERLEUKIN (IL)-6 BY MEANS OF WESTERN BLOT AND REAL-TIME QUANTITATIVE PCR (RT-PCR), RESPECTIVELY. TAKEN TOGETHER, THESE DATA SUGGEST THAT SIRT1 IN THE SPINAL CORD PLAYS AN IMPORTANT ROLE IN THE NEUROPATHIC PAIN IN THE RAT MODEL. 2018 12 1905 48 ENHANCER OF ZESTE HOMOLOG 2 CONTRIBUTES TO APOPTOSIS BY INACTIVATING JANUS KINASE 2/ SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION SIGNALING IN INFLAMMATORY BOWEL DISEASE. BACKGROUND: INFLAMMATORY BOWEL DISEASE (IBD) IS A PREVALENT WORLDWIDE HEALTH PROBLEM FEATURED BY RELAPSING, CHRONIC GASTROINTESTINAL INFLAMMATION. ENHANCER OF ZESTE HOMOLOG 2 (EZH2) IS A CRITICAL EPIGENETIC REGULATOR IN DIFFERENT PATHOLOGICAL MODELS, SUCH AS CANCER AND INFLAMMATION. HOWEVER, THE ROLE OF EZH2 IN THE IBD DEVELOPMENT IS STILL OBSCURE. AIM: TO EXPLORE THE EFFECT OF EZH2 ON IBD PROGRESSION AND THE UNDERLYING MECHANISM. METHODS: THE IBD MOUSE MODEL WAS CONDUCTED BY ADDING DEXTRAN SODIUM SULFATE (DSS), AND THE EFFECT OF EZH2 ON DSS-INDUCED COLITIS WAS ASSESSED IN THE MODEL. THE FUNCTION OF EZH2 IN REGULATING APOPTOSIS AND PERMEABILITY WAS EVALUATED BY ANNEXIN V-FITC APOPTOSIS DETECTION KIT, TRANSEPITHELIAL ELECTRICAL RESISTANCE ANALYSIS, AND WESTERN BLOT ANALYSIS OF RELATED MARKERS, INCLUDING ZONA OCCLUDENS 1, CLAUDIN-5, AND OCCLUDIN, IN NCM460 AND FETAL HUMAN COLON (FHC) CELLS. THE MECHANICAL INVESTIGATION WAS PERFORMED BY QUANTITATIVE REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION, WESTERN BLOT ANALYSIS, AND CHROMATIN IMMUNOPRECIPITATION ASSAYS. RESULTS: THE COLON LENGTH WAS INHIBITED IN THE DSS-TREATED MICE AND WAS ENHANCED BY THE EZH2 DEPLETION IN THE SYSTEM. DSS TREATMENT CAUSED A DECREASED HISTOLOGICAL SCORE IN THE MICE, WHICH WAS REVERSED BY EZH2 DEPLETION. THE INFLAMMATORY CYTOKINES, SUCH AS TUMOR NECROSIS FACTOR-ALPHA, INTERLEUKIN-6, AND INTERLEUKIN-1BETA, WERE INDUCED IN THE DSS-TREATED MICE, IN WHICH THE DEPLETION OF EZH2 COULD REVERSE THIS EFFECT. MOREOVER, THE TUMOR NECROSIS FACTOR-ALPHA TREATMENT INDUCED THE APOPTOSIS OF NCM460 AND FHC CELLS, IN WHICH EZH2 DEPLETION COULD REVERSE THIS EFFECT IN THE CELLS. MOREOVER, THE DEPLETION OF EZH2 ATTENUATED PERMEABILITY OF COLONIC EPITHELIAL CELLS. MECHANICALLY, THE DEPLETION OF EZH2 OR EZH2 INHIBITOR GSK343 WAS ABLE TO ENHANCE THE EXPRESSION AND THE PHOSPHORYLATION OF JANUS KINASE 2 (JK2) AND SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION IN THE NCM460 AND FHC CELLS. SPECIFICALLY, EZH2 INACTIVATED JAK2 EXPRESSION BY REGULATING HISTONE H3K27ME3. JAK2 INHIBITOR TG101348 WAS ABLE TO REVERSE EZH2 KNOCKDOWN-MEDIATED COLONIC EPITHELIAL CELL PERMEABILITY AND APOPTOSIS. CONCLUSION: THUS, WE CONCLUDED THAT EZH2 CONTRIBUTED TO APOPTOSIS AND INFLAMMATORY RESPONSE BY INACTIVATING JAK2/ SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION SIGNALING IN IBD. EZH2 MAY BE APPLIED AS A POTENTIAL TARGET FOR IBD THERAPY. 2021 13 3082 41 GENOME-WIDE REDISTRIBUTION OF MECP2 IN DORSAL ROOT GANGLIA AFTER PERIPHERAL NERVE INJURY. BACKGROUND: METHYL-CPG-BINDING PROTEIN 2 (MECP2), A PROTEIN WITH AFFINITY FOR METHYLATED CYTOSINES, IS CRUCIAL FOR NEURONAL DEVELOPMENT AND FUNCTION. MECP2 REGULATES GENE EXPRESSION THROUGH ACTIVATION, REPRESSION AND CHROMATIN REMODELING. MUTATIONS IN MECP2 CAUSE RETT SYNDROME, AND THESE PATIENTS DISPLAY IMPAIRED NOCICEPTION. WE OBSERVED AN INCREASE IN MECP2 EXPRESSION IN MOUSE DORSAL ROOT GANGLIA (DRG) AFTER PERIPHERAL NERVE INJURY. THE FUNCTIONAL IMPLICATION OF INCREASED MECP2 IS LARGELY UNKNOWN. TO IDENTIFY REGIONS OF THE GENOME BOUND BY MECP2 IN THE DRG AND THE CHANGES INDUCED BY NERVE INJURY, A CHROMATIN IMMUNOPRECIPITATION OF MECP2 FOLLOWED BY SEQUENCING (CHIP-SEQ) WAS PERFORMED 4 WEEKS AFTER SPARED NERVE INJURY (SNI). RESULTS: WHILE THE NUMBER OF BINDING SITES ACROSS THE GENOME REMAINED SIMILAR IN THE SNI MODEL AND SHAM CONTROL, SNI INDUCED THE REDISTRIBUTION OF MECP2 TO TRANSCRIPTIONALLY RELEVANT REGIONS. TO DETERMINE HOW DIFFERENTIAL BINDING OF MECP2 CAN AFFECT GENE EXPRESSION IN THE DRG, WE INVESTIGATED MMU-MIR-126, A MICRORNA LOCUS THAT HAD ENRICHED MECP2 BINDING IN THE SNI MODEL. ENRICHED MECP2 BINDING TO MIR-126 LOCUS AFTER NERVE INJURY REPRESSED MIR-126 EXPRESSION, AND THIS WAS NOT MEDIATED BY ALTERATIONS IN METHYLATION PATTERN AT THE MIR-126 LOCUS. DOWNREGULATION OF MIR-126 RESULTED IN THE UPREGULATION OF ITS TWO TARGET GENES DNMT1 AND VEGFA IN NEURO 2A CELLS AND IN SNI MODEL COMPARED TO CONTROL. THESE TARGET GENES WERE SIGNIFICANTLY DOWNREGULATED IN MECP2-NULL MICE COMPARED TO WILD-TYPE LITTERMATES, INDICATING A REGULATORY ROLE FOR MECP2 IN ACTIVATING DNMT1 AND VEGFA EXPRESSION. INTRATHECAL DELIVERY OF MIR-126 WAS NOT SUFFICIENT TO REVERSE NERVE INJURY-INDUCED MECHANICAL AND THERMAL HYPERSENSITIVITY, BUT DECREASED DNMT1 AND VEGFA EXPRESSION IN THE DRG. CONCLUSIONS: OUR STUDY SHOWS A REGULATORY ROLE FOR MECP2 IN THAT CHANGES IN GLOBAL REDISTRIBUTION CAN RESULT IN DIRECT AND INDIRECT MODULATION OF GENE EXPRESSION IN THE DRG. ALTERATIONS IN GENOME-WIDE BINDING OF MECP2 THEREFORE PROVIDE A MOLECULAR BASIS FOR A BETTER UNDERSTANDING OF EPIGENETIC REGULATION-INDUCED MOLECULAR CHANGES UNDERLYING NERVE INJURY. 2016 14 1945 39 EPIGALLOCATECHIN-3-GALLATE, A HISTONE ACETYLTRANSFERASE INHIBITOR, INHIBITS EBV-INDUCED B LYMPHOCYTE TRANSFORMATION VIA SUPPRESSION OF RELA ACETYLATION. BECAUSE THE P300/CBP-MEDIATED HYPERACETYLATION OF RELA (P65) IS CRITICAL FOR NUCLEAR FACTOR-KAPPAB (NF-KAPPAB) ACTIVATION, THE ATTENUATION OF P65 ACETYLATION IS A POTENTIAL MOLECULAR TARGET FOR THE PREVENTION OF CHRONIC INFLAMMATION. DURING OUR ONGOING SCREENING STUDY TO IDENTIFY NATURAL COMPOUNDS WITH HISTONE ACETYLTRANSFERASE INHIBITOR (HATI) ACTIVITY, WE IDENTIFIED EPIGALLOCATECHIN-3-GALLATE (EGCG) AS A NOVEL HATI WITH GLOBAL SPECIFICITY FOR THE MAJORITY OF HAT ENZYMES BUT WITH NO ACTIVITY TOWARD EPIGENETIC ENZYMES INCLUDING HDAC, SIRT1, AND HMTASE. AT A DOSE OF 100 MICROMOL/L, EGCG ABROGATES P300-INDUCED P65 ACETYLATION IN VITRO AND IN VIVO, INCREASES THE LEVEL OF CYTOSOLIC IKAPPABALPHA, AND SUPPRESSES TUMOR NECROSIS FACTOR ALPHA (TNFALPHA)-INDUCED NF-KAPPAB ACTIVATION. WE ALSO SHOWED THAT EGCG PREVENTS TNFALPHA-INDUCED P65 TRANSLOCATION TO THE NUCLEUS, CONFIRMING THAT HYPERACETYLATION IS CRITICAL FOR NF-KAPPAB TRANSLOCATION AS WELL AS ACTIVITY. FURTHERMORE, EGCG TREATMENT INHIBITED THE ACETYLATION OF P65 AND THE EXPRESSION OF NF-KAPPAB TARGET GENES IN RESPONSE TO DIVERSE STIMULI. FINALLY, EGCG REDUCED THE BINDING OF P300 TO THE PROMOTER REGION OF INTERLEUKIN-6 GENE WITH AN INCREASED RECRUITMENT OF HDAC3, WHICH HIGHLIGHTS THE IMPORTANCE OF THE BALANCE BETWEEN HATS AND HISTONE DEACETYLASES IN THE NF-KAPPAB-MEDIATED INFLAMMATORY SIGNALING PATHWAY. IMPORTANTLY, EGCG AT 50 MICROMOL/L DOSE COMPLETELY BLOCKS EBV INFECTION-INDUCED CYTOKINE EXPRESSION AND SUBSEQUENTLY THE EBV-INDUCED B LYMPHOCYTE TRANSFORMATION. THESE RESULTS SHOW THE CRUCIAL ROLE OF ACETYLATION IN THE DEVELOPMENT OF INFLAMMATORY-RELATED DISEASES. 2009 15 5266 40 PROMOTED INTERACTION OF C/EBPALPHA WITH DEMETHYLATED CXCR3 GENE PROMOTER CONTRIBUTES TO NEUROPATHIC PAIN IN MICE. DNA METHYLATION HAS BEEN IMPLICATED IN THE PATHOGENESIS OF CHRONIC PAIN. HOWEVER, THE SPECIFIC GENES REGULATED BY DNA METHYLATION UNDER NEUROPATHIC PAIN CONDITION REMAIN LARGELY UNKNOWN. HERE WE INVESTIGATED HOW CHEMOKINE RECEPTOR CXCR3 IS REGULATED BY DNA METHYLATION AND HOW IT CONTRIBUTES TO NEUROPATHIC PAIN INDUCED BY SPINAL NERVE LIGATION (SNL) IN MICE. SNL INCREASED CXCR3 MRNA AND PROTEIN EXPRESSION IN THE NEURONS OF THE SPINAL CORD. MEANWHILE, THE CPG (5'-CYTOSINE-PHOSPHATE-GUANINE-3') ISLAND IN THE CXCR3 GENE PROMOTER REGION WAS DEMETHYLATED, AND THE EXPRESSION OF DNA METHYLTRANSFERASE 3B (DNMT3B) WAS DECREASED. SNL ALSO INCREASED THE BINDING OF CCAAT (CYTIDINE-CYTIDINE-ADENOSINE-ADENOSINE-THYMIDINE)/ENHANCER BINDING PROTEIN ALPHA (C/EBPALPHA) WITH CXCR3 PROMOTER AND DECREASED THE BINDING OF DNMT3B WITH CXCR3 PROMOTER IN THE SPINAL CORD. C/EBPALPHA EXPRESSION WAS INCREASED IN SPINAL NEURONS AFTER SNL, AND INHIBITION OF C/EBPALPHA BY INTRATHECAL SMALL INTERFERING RNA ATTENUATED SNL-INDUCED PAIN HYPERSENSITIVITY AND REDUCED CXCR3 EXPRESSION. FURTHERMORE, SNL-INDUCED MECHANICAL ALLODYNIA AND HEAT HYPERALGESIA WERE MARKEDLY REDUCED IN CXCR3(-/-) MICE. SPINAL INHIBITION OF CXCR3 BY SHRNA OR CXCR3 ANTAGONIST ALSO ATTENUATED ESTABLISHED NEUROPATHIC PAIN. MOREOVER, CXCL10, THE LIGAND OF CXCR3, WAS INCREASED IN SPINAL NEURONS AND ASTROCYTES AFTER SNL. SUPERFUSING SPINAL CORD SLICES WITH CXCL10 ENHANCED SPONTANEOUS EPSCS AND POTENTIATED NMDA-INDUCED AND AMPA-INDUCED CURRENTS OF LAMINA II NEURONS. FINALLY, INTRATHECAL INJECTION OF CXCL10 INDUCED CXCR3-DEPENDENT PAIN HYPERSENSITIVITY IN NAIVE MICE. COLLECTIVELY, OUR RESULTS DEMONSTRATED THAT CXCR3, INCREASED BY DNA DEMETHYLATION AND THE ENHANCED INTERACTION WITH C/EBPALPHA, CAN BE ACTIVATED BY CXCL10 TO FACILITATE EXCITATORY SYNAPTIC TRANSMISSION AND CONTRIBUTE TO THE MAINTENANCE OF NEUROPATHIC PAIN. SIGNIFICANCE STATEMENT: PERIPHERAL NERVE INJURY INDUCES CHANGES OF GENE EXPRESSION IN THE SPINAL CORD THAT MAY CONTRIBUTE TO THE PATHOGENESIS OF NEUROPATHIC PAIN. CXCR3 IS A CHEMOKINE RECEPTOR. WHETHER IT IS INVOLVED IN NEUROPATHIC PAIN AND HOW IT IS REGULATED AFTER NERVE INJURY REMAIN LARGELY UNKNOWN. OUR STUDY DEMONSTRATES THAT SPINAL NERVE LIGATION DOWNREGULATES THE EXPRESSION OF DNMT3B, WHICH MAY CAUSE DEMETHYLATION OF CXCR3 GENE PROMOTER AND FACILITATE THE BINDING OF CCAAT/ENHANCER BINDING PROTEIN ALPHA WITH CXCR3 PROMOTER AND FURTHER INCREASE CXCR3 EXPRESSION IN SPINAL NEURONS. THE UPREGULATED CXCR3 MAY CONTRIBUTE TO NEUROPATHIC PAIN BY FACILITATING CENTRAL SENSITIZATION. OUR STUDY REVEALS AN EPIGENETIC MECHANISM UNDERLYING CXCR3 EXPRESSION AND ALSO SUGGESTS THAT TARGETING THE EXPRESSION OR ACTIVATION OF CXCR3 SIGNALING MAY OFFER NEW THERAPEUTICS FOR NEUROPATHIC PAIN. 2017 16 2365 42 EPIGENETIC REGULATION OF SPINAL CXCR2 SIGNALING IN INCISIONAL HYPERSENSITIVITY IN MICE. BACKGROUND: THE REGULATION OF GENE EXPRESSION IN NOCICEPTIVE PATHWAYS CONTRIBUTES TO THE INDUCTION AND MAINTENANCE OF PAIN SENSITIZATION. HISTONE ACETYLATION IS A KEY EPIGENETIC MECHANISM CONTROLLING CHROMATIN STRUCTURE AND GENE EXPRESSION. CHEMOKINE CC MOTIF RECEPTOR 2 (CXCR2) IS A PROINFLAMMATORY RECEPTOR IMPLICATED IN NEUROPATHIC AND INFLAMMATORY PAIN AND IS KNOWN TO BE REGULATED BY HISTONE ACETYLATION IN SOME SETTINGS. THE AUTHORS SOUGHT TO INVESTIGATE THE ROLE OF HISTONE ACETYLATION ON SPINAL CXCR2 SIGNALING AFTER INCISION. METHODS: GROUPS OF 5-8 MICE UNDERWENT HIND PAW INCISION. SUBEROYLANILIDE HYDROXAMIC ACID AND ANACARDIC ACID WERE USED TO INHIBIT HISTONE DEACETYLASE AND HISTONE ACETYLTRANSFERASE, RESPECTIVELY. BEHAVIORAL MEASURES OF THERMAL AND MECHANICAL SENSITIZATION AS WELL AS HYPERALGESIC PRIMING WERE USED. BOTH MESSAGE RNA QUANTIFICATION AND CHROMATIN IMMUNOPRECIPITATION ANALYSIS WERE USED TO STUDY THE REGULATION OF CXCR2 AND LIGAND EXPRESSION. FINALLY, THE SELECTIVE CXCR2 ANTAGONIST SB225002 WAS ADMINISTERED INTRATHECALLY TO REVEAL THE FUNCTION OF SPINAL CXCR2 RECEPTORS AFTER HIND PAW INCISION. RESULTS: SUBEROYLANILIDE HYDROXAMIC ACID SIGNIFICANTLY EXACERBATED MECHANICAL SENSITIZATION AFTER INCISION. CONVERSELY, ANACARDIC ACID REDUCED INCISIONAL SENSITIZATION AND ALSO ATTENUATED INCISION-INDUCED HYPERALGESIC PRIMING. OVERALL, ACETYLATED HISTONE H3 AT LYSINE 9 WAS INCREASED IN SPINAL CORD TISSUES AFTER INCISION, AND ENHANCED ASSOCIATION OF ACETYLATED HISTONE H3 AT LYSINE 9 WITH THE PROMOTER REGIONS OF CXCR2 AND KERATINOCYTE-DERIVED CHEMOKINE (CXCL1) WAS OBSERVED AS WELL. BLOCKING CXCR2 REVERSED MECHANICAL HYPERSENSITIVITY AFTER HIND PAW INCISION. CONCLUSIONS: HISTONE MODIFICATION IS AN IMPORTANT EPIGENETIC MECHANISM REGULATING INCISION-INDUCED NOCICEPTIVE SENSITIZATION. THE SPINAL CXCR2 SIGNALING PATHWAY IS ONE EPIGENETICALLY REGULATED PATHWAY CONTROLLING EARLY AND LATENT SENSITIZATION AFTER INCISION. 2013 17 4497 35 MORPHINE LEADS TO GLOBAL GENOME CHANGES IN H3K27ME3 LEVELS VIA A POLYCOMB REPRESSIVE COMPLEX 2 (PRC2) SELF-REGULATORY MECHANISM IN MESCS. BACKGROUND: ENVIRONMENTALLY INDUCED EPIGENETIC CHANGES CAN LEAD TO HEALTH PROBLEMS OR DISEASE, BUT THE MECHANISMS INVOLVED REMAIN UNCLEAR. MORPHINE CAN PASS THROUGH THE PLACENTAL BARRIER LEADING TO ABNORMAL EMBRYO DEVELOPMENT. HOWEVER, THE MECHANISM BY WHICH MORPHINE CAUSES THESE EFFECTS AND HOW THEY SOMETIMES PERSIST INTO ADULTHOOD IS NOT WELL KNOWN. TO UNRAVEL THE MORPHINE-INDUCED CHROMATIN ALTERATIONS INVOLVED IN ABERRANT EMBRYO DEVELOPMENT, WE EXPLORED THE ROLE OF THE H3K27ME3/PRC2 REPRESSIVE COMPLEX IN GENE EXPRESSION AND ITS TRANSMISSION ACROSS CELLULAR GENERATIONS IN RESPONSE TO MORPHINE. RESULTS: USING MOUSE EMBRYONIC STEM CELLS AS A MODEL SYSTEM, WE FOUND THAT CHRONIC MORPHINE TREATMENT INDUCES A GLOBAL DOWNREGULATION OF THE HISTONE MODIFICATION H3K27ME3. CONVERSELY, CHIP-SEQ SHOWED A REMARKABLE INCREASE IN H3K27ME3 LEVELS AT SPECIFIC GENOMIC SITES, PARTICULARLY PROMOTERS, DISRUPTING SELECTIVE TARGET GENES RELATED TO EMBRYO DEVELOPMENT, CELL CYCLE AND METABOLISM. THROUGH A SELF-REGULATORY MECHANISM, MORPHINE DOWNREGULATED THE TRANSCRIPTION OF PRC2 COMPONENTS RESPONSIBLE FOR H3K27ME3 BY ENRICHING HIGH H3K27ME3 LEVELS AT THE PROMOTER REGION. DOWNREGULATION OF PRC2 COMPONENTS PERSISTED FOR AT LEAST 48 H (4 CELL CYCLES) FOLLOWING MORPHINE REMOVAL, THOUGH PROMOTER H3K27ME3 LEVELS RETURNED TO CONTROL LEVELS. CONCLUSIONS: MORPHINE INDUCES TARGETING OF THE PRC2 COMPLEX TO SELECTED PROMOTERS, INCLUDING THOSE OF PRC2 COMPONENTS, LEADING TO CHARACTERISTIC CHANGES IN GENE EXPRESSION AND A GLOBAL REDUCTION IN H3K27ME3. FOLLOWING MORPHINE REMOVAL, ENHANCED PROMOTER H3K27ME3 LEVELS REVERT TO NORMAL SOONER THAN GLOBAL H3K27ME3 OR PRC2 COMPONENT TRANSCRIPT LEVELS. WE SUGGEST THAT H3K27ME3 IS INVOLVED IN INITIATING MORPHINE-INDUCED CHANGES IN GENE EXPRESSION, BUT NOT IN THEIR MAINTENANCE. MODEL OF POLYCOMB REPRESSIVE COMPLEX 2 (PRC2) AND H3K27ME3 ALTERATIONS INDUCED BY CHRONIC MORPHINE EXPOSURE. MORPHINE INDUCES H3K27ME3 ENRICHMENT AT PROMOTERS OF GENES ENCODING CORE MEMBERS OF THE PRC2 COMPLEX AND IS ASSOCIATED WITH THEIR TRANSCRIPTIONAL DOWNREGULATION. 2020 18 3175 37 H2AX PHOSPHORYLATION REGULATED BY P38 IS INVOLVED IN BIM EXPRESSION AND APOPTOSIS IN CHRONIC MYELOGENOUS LEUKEMIA CELLS INDUCED BY IMATINIB. INCREASING EVIDENCE SUGGESTS THAT HISTONE H2AX PLAYS A CRITICAL ROLE IN REGULATION OF TUMOR CELL APOPTOSIS AND ACTS AS A NOVEL HUMAN TUMOR SUPPRESSOR PROTEIN. HOWEVER, THE ACTION OF H2AX IN CHRONIC MYELOGENOUS LEUKEMIA (CML) CELLS IS UNKNOWN. THE DETAILED MECHANISM AND EPIGENETIC REGULATION BY H2AX REMAIN ELUSIVE IN CANCER CELLS. HERE, WE REPORT THAT H2AX WAS INVOLVED IN APOPTOSIS OF CML CELLS. OVEREXPRESSION OF H2AX INCREASED APOPTOTIC SENSITIVITY OF CML CELLS (K562) INDUCED BY IMATINIB. HOWEVER, OVEREXPRESSION OF SER139-MUTATED H2AX (BLOCKING PHOSPHORYLATION) DECREASED SENSITIVITY OF K562 CELLS TO APOPTOSIS. SIMILARLY, KNOCKDOWN OF H2AX MADE K562 CELLS RESISTANT TO APOPTOTIC INDUCTION. THESE RESULTS REVEALED THAT THE FUNCTION OF H2AX INVOLVED IN APOPTOSIS IS STRICTLY RELATED TO ITS PHOSPHORYLATION (SER139). OUR DATA FURTHER INDICATED THAT IMATINIB MAY STIMULATE MITOGEN-ACTIVATED PROTEIN KINASE (MAPK) FAMILY MEMBER P38, AND H2AX PHOSPHORYLATION FOLLOWED A SIMILAR TIME COURSE, SUGGESTING A PARALLEL RESPONSE. H2AX PHOSPHORYLATION CAN BE BLOCKED BY P38 SIRNA OR ITS INHIBITOR. THESE DATA DEMONSTRATED THAT H2AX PHOSPHORYLATION WAS REGULATED BY P38 MAPK PATHWAY IN K562 CELLS. HOWEVER, THE P38 MAPK DOWNSTREAM, MITOGEN- AND STRESS-ACTIVATED PROTEIN KINASE-1 AND -2, WHICH PHOSPHORYLATED HISTONE H3, WERE NOT REQUIRED FOR H2AX PHOSPHORYLATION DURING APOPTOSIS. FINALLY, WE PROVIDED EPIGENETIC EVIDENCE THAT H2AX PHOSPHORYLATION REGULATED APOPTOSIS-RELATED GENE BIM EXPRESSION. BLOCKING OF H2AX PHOSPHORYLATION INHIBITED BIM GENE EXPRESSION. TAKEN TOGETHER, THESE DATA DEMONSTRATED THAT H2AX PHOSPHORYLATION REGULATED BY P38 IS INVOLVED IN BIM EXPRESSION AND APOPTOSIS IN CML CELLS INDUCED BY IMATINIB. 2014 19 2353 31 EPIGENETIC REGULATION OF OPIOID-INDUCED HYPERALGESIA, DEPENDENCE, AND TOLERANCE IN MICE. REPEATED ADMINISTRATION OF OPIOIDS SUCH AS MORPHINE INDUCES PERSISTENT BEHAVIORAL CHANGES INCLUDING OPIOID-INDUCED HYPERALGESIA (OIH), TOLERANCE, AND PHYSICAL DEPENDENCE. IN THE CURRENT WORK WE EXPLORED HOW THE BALANCE OF HISTONE ACETYLTRANSFERASE (HAT) VERSUS HISTONE DEACETYLASE (HDAC) MIGHT REGULATE THESE MORPHINE-INDUCED CHANGES. NOCICEPTIVE THRESHOLDS, ANALGESIA, AND PHYSICAL DEPENDENCE WERE ASSESSED DURING AND FOR A PERIOD OF SEVERAL WEEKS AFTER MORPHINE EXPOSURE. TO PROBE THE ROLES OF HISTONE ACETYLATION, THE HAT INHIBITOR CURCUMIN OR A SELECTIVE HDAC INHIBITOR SUBEROYLANILIDE HYDROXAMIC ACID (SAHA) WAS ADMINISTERED DAILY TO GROUPS OF ANIMALS. HISTONE ACETYLATION IN SPINAL CORD WAS ASSESSED BY WESTERN BLOT AND IMMUNOHISTOCHEMISTRY. CONCURRENT ADMINISTRATION OF CURCUMIN WITH MORPHINE FOR 4 DAYS SIGNIFICANTLY REDUCED DEVELOPMENT OF OPIOID-INDUCED MECHANICAL ALLODYNIA, THERMAL HYPERALGESIA, TOLERANCE, AND PHYSICAL DEPENDENCE. CONVERSELY, THE HDAC INHIBITOR SAHA ENHANCED THESE RESPONSES. INTERESTINGLY, SAHA TREATMENT AFTER THE TERMINATION OF OPIOID ADMINISTRATION SUSTAINED THESE BEHAVIORAL CHANGES FOR AT LEAST 4 WEEKS. HISTONE H3 ACETYLATION IN THE DORSAL HORN OF THE SPINAL CORD WAS INCREASED AFTER CHRONIC MORPHINE TREATMENT, BUT H4 ACETYLATION WAS UNCHANGED. MOREOVER, WE OBSERVED A DECREASE IN HDAC ACTIVITY IN THE SPINAL CORDS OF MORPHINE-TREATED MICE WHILE OVERALL HAT ACTIVITY WAS UNCHANGED, SUGGESTING A SHIFT TOWARD A STATE OF ENHANCED HISTONE ACETYLATION. PERSPECTIVE: THE CURRENT STUDY INDICATES THAT EPIGENETIC MECHANISMS PLAY A CRUCIAL ROLE IN OPIOID-INDUCED LONG-LASTING NEUROPLASTICITY. THESE RESULTS PROVIDE NEW SIGHT INTO UNDERSTANDING THE MECHANISMS OF OPIOID-INDUCED NEUROPLASTICITY AND SUGGEST NEW STRATEGIES TO LIMIT OPIOID ABUSE POTENTIAL AND INCREASE THE VALUE OF THESE DRUGS AS ANALGESICS. 2013 20 2370 33 EPIGENETIC REGULATION OF THE ALTERNATIVELY ACTIVATED MACROPHAGE PHENOTYPE. ALTERNATIVELY ACTIVATED (M2) MACROPHAGES PLAY CRITICAL ROLES IN DIVERSE CHRONIC DISEASES, INCLUDING PARASITE INFECTIONS, CANCER, AND ALLERGIC RESPONSES. HOWEVER, LITTLE IS KNOWN ABOUT THE ACQUISITION AND MAINTENANCE OF THEIR PHENOTYPE. WE REPORT THAT M2-MACROPHAGE MARKER GENES ARE EPIGENETICALLY REGULATED BY RECIPROCAL CHANGES IN HISTONE H3 LYSINE-4 (H3K4) AND HISTONE H3 LYSINE-27 (H3K27) METHYLATION; AND THE LATTER METHYLATION MARKS ARE REMOVED BY THE H3K27 DEMETHYLASE JUMONJI DOMAIN CONTAINING 3 (JMJD3). WE FOUND THAT CONTINUOUS INTERLEUKIN-4 (IL-4) TREATMENT LEADS TO DECREASED H3K27 METHYLATION, AT THE PROMOTER OF M2 MARKER GENES, AND A CONCOMITANT INCREASE IN JMJD3 EXPRESSION. FURTHERMORE, WE DEMONSTRATE THAT IL-4-DEPENDENT JMJD3 EXPRESSION IS MEDIATED BY STAT6, A MAJOR TRANSCRIPTION FACTOR OF IL-4-MEDIATED SIGNALING. AFTER IL-4 STIMULATION, ACTIVATED STAT6 IS INCREASED AND BINDS TO CONSENSUS SITES AT THE JMJD3 PROMOTER. INCREASED JMJD3 CONTRIBUTES TO THE DECREASE OF H3K27 DIMETHYLATION AND TRIMETHYLATION (H3K27ME2/3) MARKS AS WELL AS THE TRANSCRIPTIONAL ACTIVATION OF SPECIFIC M2 MARKER GENES. THE DECREASE IN H3K27ME2/3 AND INCREASE IN JMJD3 RECRUITMENT WERE CONFIRMED BY IN VIVO STUDIES USING A SCHISTOSOMA MANSONI EGG-CHALLENGED MOUSE MODEL, A WELL-STUDIED SYSTEM KNOWN TO SUPPORT AN M2 PHENOTYPE. COLLECTIVELY, THESE DATA INDICATE THAT CHROMATIN REMODELING IS MECHANISTICALLY IMPORTANT IN THE ACQUISITION OF THE M2-MACROPHAGE PHENOTYPE. 2009